Identification And Characterization Of Low Temperature Induced Proteins In Chorispora Bungeana

Low temperature is one of major environmental factors that affect plant growth, development and geographical distribution. Chorispora bungeana Fisch.&C. A.Mey (C. bungeana) is a rare alpine subnival plant species that is highly capable of resisting freezing environment. Previous researches show that alpine subnival plants do not possess special morphological characteristics that help it survive under freezing environment. Thus molecular and physiological mechanisms were assumed to account for their adaptation to the freezing environment.In the present study, a comparative proteomic analysis of C. bungeana leaves was performed to identify the proteins whose expression level increased significantly in freezing stress. The results are as follows:1. Two-dimensional gel analysis combined with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) revealed that four proteins of C. bungeana which expression level increased most significantly during freezing stress were Glycine-rich RNA binding protein (GRP), DEAD-box RNA helicase (DRH), calreticulin (CRT) and Cysteine-type peptidase(CTP) respectively. Among these proteins, GRP and DRH are RNA chaperone involved in post-transcription regulation of gene expression. CRT is a multi-functional Ca2+binding protein mainly existed in endoplasmic reticulum, it plays important role in homeostasis and molecular chaperoning. CTP is a protein with unknown function.These results show that post-transcription regulation of gene expression and Ca2+signaling closely relate to low-temperature stress response in Chorispora bungeana.2. A new full-length cDNA of GRP from C. bungeana was cloned.Sequence analysis revealed that the full-length cDNA of CbGRP is1120bp. The encoding region of CbGRP is528bp. The deduced polypeptide sequence consists of176amino acid with a N-terminal RNA recognition motif (RRM) and a C-terminal glycine-rich domain containing two RGG box.3. The transcript level of CbGRP increased under low-temperature stress, salt stress and exogenous ABA. This result indicated that CbGRP is mult-stress response gene and its transcription might be regulated by ABA under stress conditions.4. Compared to non-transgenic tobacco plant, transgenic tobacco plant overexpressing CbGRP exihibits higher freezing tolerance.5. A new full-length cDNA of calreticulin (CRT) from C. bungeana was cloned. The full-length cDNA of CbCRT is1561bp with1272bp open reading region, encoding424amino acid. CbCRT has higher amino acid homology to CRT1from Arabidopsis thaliana. Amino acid sequence analysis reveals that CbCRT contain a signal peptide in its N-terminal and a putative endoplasmic reticulum targeting tetrapeptide HDEL in its C-terminal. CbCRT has two conserved calreticulin family signatures and two calreticulin family repeat motifs in its P-domain.6. The transcript level of CbCRT increased under low-temperature stress, salt stress and exogenous ABA. This result indicated that CbCRT is mult-stress response gene and its transcription might be regulated by ABA under stress conditions.